Helical-repeat dependence of integrative recombination of bacteriophage lambda: role of the P1 and H1 protein binding sites.

Abstract

The efficiency of site-specific recombination of bacteriophage lambda was found to depend on the spacing between distant protein binding sites. Insertions and deletions of up to 30 base pairs were made in the nonessential regions between the H1 and H2 protein binding sites. Recombination was found to occur in substrates with changes of integral multiples of a DNA helical repeat, whereas recombination was defective in substrates with nonintegral changes. The lambda recombinogenic complex is especially interesting because two different proteins are involved: integration host factor (IHF), which has been shown to bend DNA, and the phage-encoded integrase protein (Int), which has been shown to have two distinct DNA-binding domains. The importance of angular displacement of protein binding sites was confirmed by addition of ethidium bromide to defective substrates. Significant stimulation of recombination was observed when sufficient drug intercalated and unwound the DNA to allow improved orientation of sites. The orientation effects are dependent on supercoiling, as spacing is less important in conditions where supercoiling and the P1-H1 sites are not required for recombination.